Abstract 4131: Long-term anti-tumor immunity induced by of HPN328, a DLL3-targeting trispecific, half-life extended T cell engager, in a preclinical immunocompetent mouse model
Authors: Mary Ellen Molloy, Laura B. Valenzuela, Chi-Heng Wu, Holger Wesche, Banmeet S. Anand
Journal: Cancer Research (2023)

Abstract Tri-specific T Cell-Activating Constructs (TriTACs) are T-cell engagers that have been developed to redirect T cells to kill tumor cells. HPN328 is a Delta-like ligand 3 (DLL3)-targeting TriTAC currently being evaluated in a Phase 1/2 clinical trial enrolling patients with advanced cancers associated with DLL3 expression, including small cell lung cancer (SCLC) and other neuroendocrine malignancies (NCT04471727). HPN328 consists of three binding domains: a CD3 binder for T cell engagement, an albumin binder for half-life extension, and a DLL3 binder for tumor cell engagement. We have previously reported that HPN328 induces potent dose-dependent killing of DLL3 expressing SCLC cell lines together with dose-dependent T cell activation and cytokine release in T cell co-cultures in vitro. Based on these data and the proposed mechanism of TriTAC induced tumor cell killing we hypothesized that HPN328 could induce epitope spreading and long-term immunogenic effects. To test this hypothesis, we generated an immunocompetent mouse model in which six amino acid residues corresponding to the portion of human CD3ε which TriTAC molecules bind were added to the N terminus of the mouse CD3ε subunit. Homozygous knock-in mice (hCD3ε) were implanted subcutaneously with MC38 murine colon cancer cell line expressing human DLL3 and treated with HPN328 following tumor establishment. HPN328 treatment led to eradication of established tumors in mice. In addition, tumor infiltrating lymphocyte (TIL) analysis was performed on tumors harvested on day 4 post HPN328 treatment. Activation markers CD69 and CD25 were induced on both CD4+ and CD8+ TILs in this model. To evaluate if HPN328 can induce long-term immunity, mice whose tumors were cured by HPN328 were rechallenged on the opposite flank. As expected, tumors grew in treatment naïve mice, but no tumors were detected in mice previously treated with HPN328, demonstrating sustained and durable anti-tumor immunity induced by HPN328 treatment. Anti-tumor immunity is an indicator of T cell memory against tumor antigens. Concomitant with anti-tumor immune response, we detected increases in memory T cells in the spleens of mice previously treated with HPN328. Together these results indicate that HPN328 can induce epitope spreading and prolonged anti-tumor immunity, suggest a novel mechanism for its activity and efficacy in vivo. Citation Format: Mary Ellen Molloy, Laura B. Valenzuela, Chi-Heng Wu, Holger Wesche, Banmeet S. Anand. Long-term anti-tumor immunity induced by of HPN328, a DLL3-targeting trispecific, half-life extended T cell engager, in a preclinical immunocompetent mouse model. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4131.